环形泰勒虫转化细胞表达外源基因系统的构建OA

To improve the transfection efficiency,this study used TaNM1 cells transformed by thei-leria annulata as the research target,and obtained RNA encoding enhanced green fluorescent protein(eGFP)protein by in vitro transcription(IVT)technology.After capping and purification,eGFP mRNA was transfected into TaNM1 cells by liposomes.Blank cells was set as negative control,while cells trans-fected with p-eGFP plasmid by either liposome or electro-transfection was set as positive control.The expression of eGFP protein in TaNM1 cells was detected by Western-blot,fluorescence microscope and flow cytometry.The results showed that green fluorescence could be observed when eGFP mRNA was trans-fected into TaNM1 cells with liposomes,and the expression of eGFP protein(27 kDa)was also detected by Western-blot,and the transfection efficiency could reach 66.22％by flow cytometry.The p-eGFP plasmid by liposome or electro-transfection could not induce the expression of eGFP in TaNM1 cells,and elec-tro-transfection induced massive cell death.In conclusion,an efficient in vitro transcription system was successfully established,providing a technical support for in-depth research on the infection and pathogenic mechanism of Theileria annulata.